By David N. Leff
Hyper-macho men sneeringly dismiss gonorrhea as "no worse than a bad cold."
In fact, this reportedly most prevalent sexually transmitted disease on earth, spread by the bacterium Neisseria gonorrhoeae can cause - in men - a painful inflammation of the genital organs, and in newborn infants, blindness. Most infected women go for weeks or months oblivious to a case of gonorrhea, which in females is initially asymptomatic.
Penicillin used to be the magic bullet that cured a case of gonorrhea, but the wily N. gonorrhoeae pathogen quickly mounted resistance to this and other standard antibiotics. Nowadays, physicians treat a patient with ceftriaxone, a cephalosporin analog.
Years ago, immunologists sought to make a vaccine against the pili, or filaments, that stud the surface of Neisseria. Largely composed of an antigenic protein called pilin, these projections seemed like an ideal antibody target, not only against the gonococcus but against its deadlier counterpart, N. meningitidis, which causes life-threatening meningitis.
"What made that effort seem logical," explained microbiologist Matthew Wolfgang, at the University of Michigan, Ann Arbor, was the fact that both pathogens shared the same type of pilus structure and target proteins. When you have an infection of N. gonorrhoeae or N. meningitidis, the patient usually has an immune response that's directed against the pili.
"But 15 or 20 years ago," he recalled, "they realized that those Neisseria bacteria can rapidly change the antigenicity of their pilus subunit, and the structure became completely different. It can evade the human immune system, just like the influenza virus and HIV do. That fact brought the idea of making a vaccine against pili to a screeching halt, because these things could probably come up with an infinitely varied number of structures. So, my colleagues' objective then became to understand how does the pathogen make pili. Because pili are absolutely essential for infecting the human host. So, let's figure out how these structures are assembled and how they work, which maybe would give us some better targets to develop vaccines, or go after therapeutic drugs."
Twitching Motility Spreads Disease
A graduate student in the university department of microbiology and immunology, Wolfgang is first author of a paper in today's Proceedings of the National Academy of Sciences (PNAS), dated Dec. 8, 1998. It brings their findings up to date in a study titled "Suppression of an absolute defect in Type IV pilus biogenesis by loss-of-function mutations in pilT, a twitching motility gene in Neisseria gonorrhoeae." Its senior author is microbiologist Michael Koomey.
N. gonorrhoeae is a diplococcus, that is, a double-cell bacterium, joined together like Siamese twins. "Each cell, 0.6 to 1.0 microns in diameter, is decorated with an estimated 50 pili," Wolfgang told BioWorld Today, "ranging in length from 1,000 to 4,000 nanometers, by 60 Angstroms in diameter. We refer to the pilus as a surface organelle," he observed. "It mediates adherence of the bacterium to its human host." (See also BioWorld Today, Nov. 20, 1998, p. 1.)
"N. gonorrhea's sole habitat," Wolfgang pointed out, "is human epithelial cells, where it grows naturally - but not in any other mammals. It adheres very robustly to corneal epithelial cells, and is a naturally occurring site of infection is the eye. Gonococcal eye infections are a problem with newborns; it can result in blindness."
The PNAS paper reports testing in donated human corneas of wild-type N. gonorrhoeae cells and mutants that the co-authors have created.
"They infected a monolayer of human corneal cells," Wolfgang recounted, "with the bacterial cells. Then we could assess the ability of those bacteria to adhere to those epithelial cells."
Wild-type N. gonorrhoeae cells have two main activities in life: assemble and disassemble pili; grab onto passing DNA.
"Type IV pili are a little unique," Wolfgang explained. "They correlate with the ability of the bacteria to take up DNA from the environment, a process called natural transformation. But, in order to take up DNA, the bacteria have to express Type IV pili. Neisseria spontaneously may lyse and spill out their DNA. The cells break open at reasonably high frequency. Usually, when their population reaches a high density, a certain fraction will autolyse, and spill their DNA into the environment."
Gonorrhea Cells Give Each Other Gonorrhea
"Other gonococcal cells present in the population are capable of taking up that DNA. This looks like cannibalism, but it's more likely a form of bacterial sex," Wolfgang continued.
"These Type IV pili also play a role in twitching motility. This is a non-flagellar, jerky form of motility, in which the bacteria vibrate, and maybe spread out. Of course, no one really understands it. Several models propose that once the pathogen adheres to its human host in localized fashion, as a big clump of bacteria, then it uses twitching motility to disperse this clump and disseminate the infection."
Elucidating the N. gonorrhoeae pilus's life cycle made sense of the organelle's quick-change artistry, which frustrated those pioneer vaccinologists.
"In Neisseria," Wolfgang said, "they expressed the pilin protein - the major structural subunit. But these also have a repertoire of 15 to 20 silent copies of the pilin gene, called pilS. These don't contain all the information to make the protein.
"So, Neisseria undergoes the process of gene conversion," he went on. They can swap in fragments of these pilS genes into the pilin gene, which is expressed. The end product is antigenic variation. And the variable epitopes are probably lining the outside of the pilus. So, by swapping alleles into expression genes you can make a completely different subunit, at least antigenically different, so it looks completely strange to the host immune system.
"Our particular objective now," Wolfgang concluded, "is to make this information available, so maybe other groups can use it to try either to make vaccines that go after other components required for masking the pili, or hunt for compounds that block pili assembly." N